It is important to measure and assess the role of humoral immune responses elicited by prophylactic HIV vaccine regimens. In particular, we have focused on the development of DNA vaccine technology as a putative prophylactic vaccine for HIV-1. We have established that these vaccines are capable of eliciting protection in some primate model systems under some circumstances. In addition, these vaccines have progressed to the clinic for preliminary phase I trials of safety and immunogenicity. Analysis of a completed therapeutic phase I trial indicated little acute safety concerns and demonstrated that boosting of both anti-gp120 humoral immune responses as well as enhanced cellular (i.e. cytotoxic T cell responses) responses could be achieved. Humoral immune responses, however, in HIV-1 negative chimpanzees and macaques have been of modest titer. It is the overall goal of this NCVDG to improve on these initial results. Therefore, the development of methods which will enhance humoral immune responses to these DNA vaccine preparations are of importance. In this regard "Molecular Adjuvants" for DNA vaccines such as cytokine gene sequences have been shown to enhance both humoral and cellular immune responses in vivo. The major purpose of this proposal is to measure two specific immunological variables following co-administration of a battery of "molecular" vaccine adjuvants along with well characterized DNA plasmid based HIV vaccine constructs. Specifically, these immunological parameters are : (a) measurement of specific functional antibody responses in rodents, non-human primates and humans and (b) to measure the levels of the possibly HIV-1 suppressive beta chemokines in vivo and in vitro from vaccinated experimental animals and humans. The rationale for measuring beta chemokines is based upon some preliminary findings which showed elevated serum levels of the beta chemokine MIP 1alpha in HIV-1 asymptomatic patients after their vaccination with a gp160 expressing DNA construct. The basic hypothesis being addressed in this application is that co-administration of selected DNA plasmids expressing cytokine-co- stimulatory molecules will functionally enhance the humoral immunogenicity and efficacy of plasmid based DNA vaccines. The data generated by these studies could have major relevance for the further development of clinically efficacious HIV-1 DNA plasmid based vaccines.